bims-mibica Biomed News
on Mitochondrial bioenergetics in cancer
Issue of 2023‒06‒11
35 papers selected by
Kelsey Fisher-Wellman
East Carolina University

  1. Cancer Res. 2023 Jun 05. pii: CAN-23-1006. [Epub ahead of print]
      Dependency on mitochondrial oxidative phosphorylation (OxPhos) is a potential weakness for leukemic stem cells (LSCs) that can be exploited for therapeutic purposes. Fatty acid oxidation (FAO) is a crucial OxPhos-fueling catabolic pathway for some acute myeloid leukemia (AML) cells, particularly chemotherapy-resistant AML cells. Here, we identified cold sensitivity at 4°C (cold killing challenge; CKC4), commonly used for sample storage, as a novel vulnerability that selectively kills AML LSCs with active FAO-supported OxPhos while sparing normal hematopoietic stem cells (HSCs). Cell death of OxPhos-positive leukemic cells was induced by membrane permeabilization at 4°C; by sharp contrast, leukemic cells relying on glycolysis were resistant. Forcing glycolytic cells to activate OxPhos metabolism sensitized them to CKC4. Lipidomic and proteomic analyzes showed that OxPhos shapes the composition of the plasma membrane and introduces variation of 22 lipid subfamilies between cold-sensitive and cold-resistant cells. Together, these findings indicate that steady-state energy metabolism at body temperature predetermines the sensitivity of AML LSCs to cold temperature, suggesting that cold sensitivity could be a potential OxPhos biomarker. These results could have important implications for designing experiments for AML research to avoid cell storage at 4°C.
  2. Nature. 2023 Jun 07.
      The mitochondrial unfolded protein response (UPRmt) is essential to safeguard mitochondria from proteotoxic damage by activating a dedicated transcriptional response in the nucleus to restore proteostasis1,2. Yet, it remains unclear how the information on mitochondria misfolding stress (MMS) is signalled to the nucleus as part of the human UPRmt (refs. 3,4). Here, we show that UPRmt signalling is driven by the release of two individual signals in the cytosol-mitochondrial reactive oxygen species (mtROS) and accumulation of mitochondrial protein precursors in the cytosol (c-mtProt). Combining proteomics and genetic approaches, we identified that MMS causes the release of mtROS into the cytosol. In parallel, MMS leads to mitochondrial protein import defects causing c-mtProt accumulation. Both signals integrate to activate the UPRmt; released mtROS oxidize the cytosolic HSP40 protein DNAJA1, which leads to enhanced recruitment of cytosolic HSP70 to c-mtProt. Consequently, HSP70 releases HSF1, which translocates to the nucleus and activates transcription of UPRmt genes. Together, we identify a highly controlled cytosolic surveillance mechanism that integrates independent mitochondrial stress signals to initiate the UPRmt. These observations reveal a link between mitochondrial and cytosolic proteostasis and provide molecular insight into UPRmt signalling in human cells.
  3. Proc Natl Acad Sci U S A. 2023 Jun 13. 120(24): e2213241120
      The inner mitochondrial membrane (IMM), housing components of the electron transport chain (ETC), is the site for respiration. The ETC relies on mobile carriers; therefore, it has long been argued that the fluidity of the densely packed IMM can potentially influence ETC flux and cell physiology. However, it is unclear if cells temporally modulate IMM fluidity upon metabolic or other stimulation. Using a photostable, red-shifted, cell-permeable molecular-rotor, Mitorotor-1, we present a multiplexed approach for quantitatively mapping IMM fluidity in living cells. This reveals IMM fluidity to be linked to cellular-respiration and responsive to stimuli. Multiple approaches combining in vitro experiments and live-cell fluorescence (FLIM) lifetime imaging microscopy (FLIM) show Mitorotor-1 to robustly report IMM 'microviscosity'/fluidity through changes in molecular free volume. Interestingly, external osmotic stimuli cause controlled swelling/compaction of mitochondria, thereby revealing a graded Mitorotor-1 response to IMM microviscosity. Lateral diffusion measurements of IMM correlate with microviscosity reported via Mitorotor-1 FLIM-lifetime, showing convergence of independent approaches for measuring IMM local-order. Mitorotor-1 FLIM reveals mitochondrial heterogeneity in IMM fluidity; between-and-within cells and across single mitochondrion. Multiplexed FLIM lifetime imaging of Mitorotor-1 and NADH autofluorescence reveals that IMM fluidity positively correlates with respiration, across individual cells. Remarkably, we find that stimulating respiration, through nutrient deprivation or chemically, also leads to increase in IMM fluidity. These data suggest that modulating IMM fluidity supports enhanced respiratory flux. Our study presents a robust method for measuring IMM fluidity and suggests a dynamic regulatory paradigm of modulating IMM local order on changing metabolic demand.
    Keywords:  fluidity; fluorescence lifetime; fluorescent probe; metabolism; mitochondria
  4. EMBO Rep. 2023 Jun 06. e57127
      The mitochondrial ADP/ATP carrier (SLC25A4), also called the adenine nucleotide translocase, imports ADP into the mitochondrial matrix and exports ATP, which are key steps in oxidative phosphorylation. Historically, the carrier was thought to form a homodimer and to operate by a sequential kinetic mechanism, which involves the formation of a ternary complex with the two exchanged substrates bound simultaneously. However, recent structural and functional data have demonstrated that the mitochondrial ADP/ATP carrier works as a monomer and has a single substrate binding site, which cannot be reconciled with a sequential kinetic mechanism. Here, we study the kinetic properties of the human mitochondrial ADP/ATP carrier by using proteoliposomes and transport robotics. We show that the Km/Vmax ratio is constant for all of the measured internal concentrations. Thus, in contrast to earlier claims, we conclude that the carrier operates with a ping-pong kinetic mechanism in which substrate exchange across the membrane occurs consecutively rather than simultaneously. These data unite the kinetic and structural models, showing that the carrier operates with an alternating access mechanism.
    Keywords:  ADP/ATP translocase; SLC25; adenine nucleotide translocator; bioenergetics; mitochondrial carrier family
  5. Proc Natl Acad Sci U S A. 2023 Jun 13. 120(24): e2216310120
      Many types of differentiated cells can reenter the cell cycle upon injury or stress. The underlying mechanisms are still poorly understood. Here, we investigated how quiescent cells are reactivated using a zebrafish model, in which a population of differentiated epithelial cells are reactivated under a physiological context. A robust and sustained increase in mitochondrial membrane potential was observed in the reactivated cells. Genetic and pharmacological perturbations show that elevated mitochondrial metabolism and ATP synthesis are critical for cell reactivation. Further analyses showed that elevated mitochondrial metabolism increases mitochondrial ROS levels, which induces Sgk1 expression in the mitochondria. Genetic deletion and inhibition of Sgk1 in zebrafish abolished epithelial cell reactivation. Similarly, ROS-dependent mitochondrial expression of SGK1 promotes S phase entry in human breast cancer cells. Mechanistically, SGK1 coordinates mitochondrial activity with ATP synthesis by phosphorylating F1Fo-ATP synthase. These findings suggest a conserved intramitochondrial signaling loop regulating epithelial cell renewal.
    Keywords:  F1Fo-ATP synthase; IGF/insulin signaling; mitochondrial membrane potential; reactive oxygen species; serum- and glucocorticoid-regulated kinase 1
  6. Mol Med. 2023 06 06. 29(1): 72
      BACKGROUND: Mitochondrial metabolism has been proposed as an attractive target for breast cancer therapy. The discovery of new mechanisms underlying mitochondrial dysfunction will facilitate the development of new metabolic inhibitors to improve the clinical treatment of breast cancer patients. DYNLT1 (Dynein Light Chain Tctex-Type 1) is a key component of the motor complex that transports cellular cargo along microtubules in the cell, but whether and how DYNLT1 affects mitochondrial metabolism and breast cancer has not been reported.METHODS: The expression levels of DYNLT1 were analyzed in clinical samples and a panel of cell lines. The role of DYNLT1 in breast cancer development was investigated using in vivo mouse models and in vitro cell assays, including CCK-8, plate cloning and transwell assay. The role of DYNLT1 in regulating mitochondrial metabolism in breast cancer development is examined by measuring mitochondrial membrane potential and ATP levels. To investigate the underlying molecular mechanism, many methods, including but not limited to Co-IP and ubiquitination assay were used.
    RESULTS: First, we found that DYNLT1 was upregulated in breast tumors, especially in ER + and TNBC subtypes. DYNLT1 promotes the proliferation, migration, invasion and mitochondrial metabolism in breast cancer cells in vitro and breast tumor development in vivo. DYNLT1 colocalizes with voltage-dependent anion channel 1 (VDAC1) on mitochondria to regulate key metabolic and energy functions. Mechanistically, DYNLT1 stabilizes the voltage-dependent anion channel 1 (VDAC1) by hindering E3 ligase Parkin-mediated VDAC1 ubiquitination and degradation.
    CONCLUSION: Our data demonstrate that DYNLT1 promotes mitochondrial metabolism to fuel breast cancer development by inhibiting Parkin-mediated ubiquitination degradation of VDAC1. This study suggests that mitochondrial metabolism can be exploited by targeting the DYNLT1-Parkin-VDAC1 axis to improve the ability of metabolic inhibitors to suppress cancers with limited treatment options, such as triple-negative breast cancer (TNBC).
    Keywords:  Breast cancer; DYNLT1; Mitochondrial metabolism; Protein stability; VDAC1
  7. Nat Cell Biol. 2023 Jun 08.
      De novo pyrimidine biosynthesis is achieved by cytosolic carbamoyl-phosphate synthetase II, aspartate transcarbamylase and dihydroorotase (CAD) and uridine 5'-monophosphate synthase (UMPS), and mitochondrial dihydroorotate dehydrogenase (DHODH). However, how these enzymes are orchestrated remains enigmatical. Here we show that cytosolic glutamate oxaloacetate transaminase 1 clusters with CAD and UMPS, and this complex then connects with DHODH, which is mediated by the mitochondrial outer membrane protein voltage-dependent anion-selective channel protein 3. Therefore, these proteins form a multi-enzyme complex, named 'pyrimidinosome', involving AMP-activated protein kinase (AMPK) as a regulator. Activated AMPK dissociates from the complex to enhance pyrimidinosome assembly but inactivated UMPS, which promotes DHODH-mediated ferroptosis defence. Meanwhile, cancer cells with lower expression of AMPK are more reliant on pyrimidinosome-mediated UMP biosynthesis and more vulnerable to its inhibition. Our findings reveal the role of pyrimidinosome in regulating pyrimidine flux and ferroptosis, and suggest a pharmaceutical strategy of targeting pyrimidinosome in cancer treatment.
  8. Life Sci. 2023 Jun 02. pii: S0024-3205(23)00451-4. [Epub ahead of print]327 121817
      AIMS: Pyruvate carboxylase (PC) plays a key role in cancer cell metabolic reprogramming. Whether metabolic reprogramming and PC are related in PDAC is unclear. Here, the effect of PC expression on PDAC tumorigenesis and metabolic reprogramming were evaluated.MATERIALS AND METHODS: PC protein expression in PDAC and precancerous tissues was measured through immunohistochemistry. The maximum standardized uptake (SUVmax) of 18F-fluoro-2-deoxy-2-d-glucose (18F-FDG) in PDAC patient PET/CT scans before surgical resection was retrospectively determined. Stable PC-knockdown and PC-overexpressing cells were established using lentiviruses, and PDAC progression was assessed in vivo and in vitro. Lactate content, 18F-FDG cell uptake rate, mitochondrial oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) were measured in cells. RNA sequencing revealed and qPCR verified differentially expressed genes (DEGs) after PC knockdown. The signaling pathways involved were determined by Western blotting.
    KEY FINDINGS: PC was significantly upregulated in PDAC tissues vs. precancerous tissues. A high SUVmax correlated with PC upregulation. PC knockdown significantly inhibited PDAC progression. Lactate content, SUVmax, and ECAR significantly decreased after PC knockdown. Peroxisome proliferator-activated receptor gamma coactivator-one alpha (PGC-1α) was upregulated after PC knockdown; and PGC1a expression promoted AMPK phosphorylation to activate mitochondrial metabolism. Metformin significantly inhibited mitochondrial respiration after PC knockdown, further activated AMPK and downstream carnitine palmitoyltransferase 1A (CPT1A)-regulated fatty acid oxidation (FAO), and inhibited PDAC cells progression.
    SIGNIFICANCE: PDAC cell uptake of FDG was positively correlated with PC expression. PC promotes PDAC glycolysis, and reducing PC expression can increase PGC1a expression, activate AMPK, and restore metformin sensitivity.
    Keywords:  FDG uptake; Metformin; OXPHOS; PC; PDAC; PGC-1α
  9. Cell Rep. 2023 Jun 07. pii: S2211-1247(23)00627-7. [Epub ahead of print]42(6): 112616
      Combined inhibition of oxidative phosphorylation (OXPHOS) and glycolysis has been shown to activate a PP2A-dependent signaling pathway, leading to tumor cell death. Here, we analyze highly selective mitochondrial complex I or III inhibitors in vitro and in vivo to elucidate the molecular mechanisms leading to cell death following OXPHOS inhibition. We show that IACS-010759 treatment (complex I inhibitor) induces a reactive oxygen species (ROS)-dependent dissociation of CIP2A from PP2A, leading to its destabilization and degradation through chaperone-mediated autophagy. Mitochondrial complex III inhibition has analogous effects. We establish that activation of the PP2A holoenzyme containing B56δ regulatory subunit selectively mediates tumor cell death, while the arrest in proliferation that is observed upon IACS-010759 treatment does not depend on the PP2A-B56δ complex. These studies provide a molecular characterization of the events subsequent to the alteration of critical bioenergetic pathways and help to refine clinical studies aimed to exploit metabolic vulnerabilities of tumor cells.
    Keywords:  CIP2A; CP: Cancer; CP: Molecular biology; OXPHOS; PP2A; cancer; chaperone-mediated autophagy; fasting; glycolysis; metabolism
  10. Phys Biol. 2023 Jun 08.
      Mitochondria serve a wide range of functions within cells, most notably via their production of ATP. Although their morphology is commonly described as bean-like, mitochondria often form interconnected networks within cells that exhibit dynamic restructuring through a variety of physical changes. Further, though relationships between form and function in biology are well established, the extant toolkit for understanding mitochondrial morphology is limited. Here, we emphasize new and established methods for quantitatively describing mitochondrial networks, ranging from unweighted graph-theoretic representations to multi-scale approaches from applied topology, in particular persistent homology. We also show fundamental relationships between mitochondrial networks, mathematics, and physics, using ideas of graph planarity and statistical mechanics to better understand the full possible morphological
space of mitochondrial network structures. Lastly, we provide suggestions for how examination of mitochondrial network form through the language of mathematics can inform biological understanding, and vice versa.
    Keywords:  graph theory; mitochondrial networks; persistent homology; planar graphs; scaling
  11. Mol Cancer Ther. 2023 Jun 08. pii: MCT-23-0149. [Epub ahead of print]
      Identifying novel, unique, and personalized molecular targets for patients with pancreatic ductal adenocarcinoma (PDAC) remains the greatest challenge in altering the biology of fatal tumors. Bromo- and Extra-Terminal domain (BET) proteins are activated in a non-canonical fashion by TGF-β, a ubiquitous cytokine in the PDAC tumor microenvironment (TME). We hypothesized that BET inhibitors (BETi) represent a new class of drugs that attack PDAC tumors via a novel mechanism. Using a combination of patient and syngeneic murine models, we investigated the effects of the BETi drug BMS-986158 on cellular proliferation, organoid growth, cell cycle progression, and mitochondrial metabolic disruption. These were investigated independently and in combination with standard cytotoxic chemotherapy (gemcitabine + paclitaxel [GemPTX] ). BMS-986158 reduced cell viability and proliferation across multiple PDAC cell lines in a dose-dependent manner, even more so in combination with cytotoxic chemotherapy (p<0.0001). We found that BMS-986158 reduced both human and murine PDAC organoid growth (p<0.001), with associated perturbations in the cell cycle leading to cell cycle arrest. BMS-986158 disrupts normal cancer-dependent mitochondrial function, leading to aberrant mitochondrial metabolism and stress via dysfunctional cellular respiration, proton leakage, and ATP production. We demonstrated mechanistic and functional data that BETi induces metabolic mitochondrial dysfunction, abrogating PDAC progression and proliferation, alone and in combination with systemic cytotoxic chemotherapies. This novel approach improves the therapeutic window in patients with PDAC and offers another treatment approach distinct from cytotoxic chemotherapy that targets cancer cell bioenergetics.
  12. ACS Nano. 2023 Jun 08.
      We present the super-resolution microscopy of functional, isolated functional mitochondria, enabling real-time studies of structure and function (voltages) in response to pharmacological manipulation. Changes in mitochondrial membrane potential as a function of time and position can be imaged in different metabolic states (not possible in whole cells), created by the addition of substrates and inhibitors of the electron transport chain, enabled by the isolation of vital mitochondria. By careful analysis of structure dyes and voltage dyes (lipophilic cations), we demonstrate that most of the fluorescent signal seen from voltage dyes is due to membrane bound dyes, and develop a model for the membrane potential dependence of the fluorescence contrast for the case of super-resolution imaging, and how it relates to membrane potential. This permits direct analysis of mitochondrial structure and function (voltage) of isolated, individual mitochondria as well as submitochondrial structures in the functional, intact state, a major advance in super-resolution studies of living organelles.
    Keywords:  Voltage; electrophysiology; fluorescent dye; lipid bilayer; metabolism; mitochondria; super-resolution
  13. Nat Commun. 2023 Jun 05. 14(1): 3251
      While targeted treatment against BRAF(V600E) improve survival for melanoma patients, many will see their cancer recur. Here we provide data indicating that epigenetic suppression of PGC1α defines an aggressive subset of chronic BRAF-inhibitor treated melanomas. A metabolism-centered pharmacological screen further identifies statins (HMGCR inhibitors) as a collateral vulnerability within PGC1α-suppressed BRAF-inhibitor resistant melanomas. Lower PGC1α levels mechanistically causes reduced RAB6B and RAB27A expression, whereby their combined re-expression reverses statin vulnerability. BRAF-inhibitor resistant cells with reduced PGC1α have increased integrin-FAK signaling and improved extracellular matrix detached survival cues that helps explain their increased metastatic ability. Statin treatment blocks cell growth by lowering RAB6B and RAB27A prenylation that reduces their membrane association and affects integrin localization and downstream signaling required for growth. These results suggest that chronic adaptation to BRAF-targeted treatments drive novel collateral metabolic vulnerabilities, and that HMGCR inhibitors may offer a strategy to treat melanomas recurring with suppressed PGC1α expression.
  14. FEBS Lett. 2023 Jun 05.
      Mitochondria are organelles indispensable for the correct functioning of eukaryotic cells. Their significance for cellular homeostasis is manifested by the existence of complex quality control pathways that monitor organellar fitness. Mitochondrial biogenesis relies on the efficient import of mitochondrial precursor proteins, a large majority of which are encoded by nuclear DNA and synthesized in the cytosol. This creates a demand for highly specialized import routes that comprise cytosolic factors and organellar translocases. The passage of newly encoded mitochondrial precursor proteins through the cytosol to the translocase of the outer mitochondrial membrane (TOM) is under tight surveillance. As a result of mitochondrial import defects, mitochondrial precursor proteins accumulate in the cytosol or clog the TOM complex, which in turn stimulates cellular stress responses to minimize the consequences of these challenges. These responses are critical for maintaining protein homeostasis under conditions of mitochondrial stress. The present review summarizes recent advances in the field of mitochondrial protein import quality control and discusses the role of this quality control within the network of cellular mechanisms that maintain the cellular homeostasis of proteins.
    Keywords:  cellular stress responses; mitochondria; mitochondrial dysfunction; mitochondrial quality control; protein aggregates; protein homeostasis
  15. Elife. 2023 Jun 05. pii: e84204. [Epub ahead of print]12
      Mitochondrial ATP production in cardiac ventricular myocytes must be continually adjusted to rapidly replenish the ATP consumed by the working heart. Two systems are known to be critical in this regulation: mitochondrial matrix Ca2+ ([Ca2+]m) and blood flow that is tuned by local ventricular myocyte metabolic signaling. However, these two regulatory systems do not fully account for the physiological range of ATP consumption observed. We report here on the identity, location, and signaling cascade of a third regulatory system -- CO2/bicarbonate. CO2 is generated in the mitochondrial matrix as a metabolic waste product of the oxidation of nutrients that powers ATP production. It is a lipid soluble gas that rapidly permeates the inner mitochondrial membrane (IMM) and produces bicarbonate (HCO3-) in a reaction accelerated by carbonic anhydrase (CA). The bicarbonate level is tracked physiologically by a bicarbonate-activated adenylyl cyclase, soluble adenylyl cyclase (sAC). Using structural Airyscan super-resolution imaging and functional measurements we find that sAC is primarily inside the mitochondria of ventricular myocytes where it generates cAMP when activated by HCO3-. Our data strongly suggest that ATP production in these mitochondria is regulated by this cAMP signaling cascade operating within the inter-membrane space (IMS) by activating local EPAC1 (Exchange Protein directly Activated by cAMP) which turns on Rap1 (Ras-related protein 1). Thus, mitochondrial ATP production is shown to be increased by bicarbonate-triggered sAC signaling through Rap1. Additional evidence is presented indicating that the cAMP signaling itself does not occur directly in the matrix. We also show that this third signaling process involving bicarbonate and sAC activates the cardiac mitochondrial ATP production machinery by working independently of, yet in conjunction with, [Ca2+]m-dependent ATP production to meet the energy needs of cellular activity in both health and disease. We propose that the bicarbonate and calcium signaling arms function in a resonant or complementary manner to match mitochondrial ATP production to the full range of energy consumption in cardiac ventricular myocytes in health and disease.
    Keywords:  biochemistry; chemical biology; molecular biophysics; rat; structural biology
  16. RSC Chem Biol. 2023 Jun 07. 4(6): 386-398
      Complex I is an essential membrane protein in respiration, oxidising NADH and reducing ubiquinone to contribute to the proton-motive force that powers ATP synthesis. Liposomes provide an attractive platform to investigate complex I in a phospholipid membrane with the native hydrophobic ubiquinone substrate and proton transport across the membrane, but without convoluting contributions from other proteins present in the native mitochondrial inner membrane. Here, we use dynamic and electrophoretic light scattering techniques (DLS and ELS) to show how physical parameters, in particular the zeta potential (ζ-potential), correlate strongly with the biochemical functionality of complex I-containing proteoliposomes. We find that cardiolipin plays a crucial role in the reconstitution and functioning of complex I and that, as a highly charged lipid, it acts as a sensitive reporter on the biochemical competence of proteoliposomes in ELS measurements. We show that the change in ζ-potential between liposomes and proteoliposomes correlates linearly with protein retention and catalytic oxidoreduction activity of complex I. These correlations are dependent on the presence of cardiolipin, but are otherwise independent of the liposome lipid composition. Moreover, changes in the ζ-potential are sensitive to the proton motive force established upon proton pumping by complex I, thereby constituting a complementary technique to established biochemical assays. ELS measurements may thus serve as a more widely useful tool to investigate membrane proteins in lipid systems, especially those that contain charged lipids.
  17. Commun Biol. 2023 Jun 08. 6(1): 618
      Mitochondria are dynamic organelles that are important for cell growth and proliferation. Dysregulated mitochondrial dynamics are highly associated with the initiation and progression of various cancers, including ovarian cancer. However, the regulatory mechanism underlying mitochondrial dynamics is still not fully understood. Previously, our study showed that carnitine palmitoyltransferase 1A (CPT1A) is highly expressed in ovarian cancer cells and promotes the development of ovarian cancer. Here, we find that CPT1A regulates mitochondrial dynamics and promotes mitochondrial fission in ovarian cancer cells. Our study futher shows that CPT1A regulates mitochondrial fission and function through mitochondrial fission factor (MFF) to promote the growth and proliferation of ovarian cancer cells. Mechanistically, we show that CPT1A promotes succinylation of MFF at lysine 302 (K302), which protects against Parkin-mediated ubiquitin-proteasomal degradation of MFF. Finally, the study shows that MFF is highly expressed in ovarian cancer cells and that high MFF expression is associated with poor prognosis in patients with ovarian cancer. MFF inhibition significantly inhibits the progression of ovarian cancer in vivo. Overall, CPT1A regulates mitochondrial dynamics through MFF succinylation to promote the development of ovarian cancer. Moreover, our findings suggest that MFF is a potential therapeutic target for ovarian cancer.
  18. bioRxiv. 2023 May 22. pii: 2023.05.22.541833. [Epub ahead of print]
      The developing mammalian heart undergoes an important metabolic shift from glycolysis toward mitochondrial oxidation, such that oxidative phosphorylation defects may present with cardiac abnormalities. Here, we describe a new mechanistic link between mitochondria and cardiac morphogenesis, uncovered by studying mice with systemic loss of the mitochondrial citrate carrier SLC25A1. Slc25a1 null embryos displayed impaired growth, cardiac malformations, and aberrant mitochondrial function. Importantly, Slc25a1 haploinsufficient embryos, which are overtly indistinguishable from wild type, exhibited an increased frequency of these defects, suggesting Slc25a1 dose-dependent effects. Supporting clinical relevance, we found a near-significant association between ultrarare human pathogenic SLC25A1 variants and pediatric congenital heart disease. Mechanistically, SLC25A1 may link mitochondria to transcriptional regulation of metabolism through epigenetic control of PPARγ to promote metabolic remodeling in the developing heart. Collectively, this work positions SLC25A1 as a novel mitochondrial regulator of ventricular morphogenesis and cardiac metabolic maturation and suggests a role in congenital heart disease.
  19. Dev Cell. 2023 Jun 02. pii: S1534-5807(23)00239-3. [Epub ahead of print]
      Cells adjust their metabolism by remodeling membrane contact sites that channel metabolites to different fates. Lipid droplet (LD)-mitochondria contacts change in response to fasting, cold exposure, and exercise. However, their function and mechanism of formation have remained controversial. We focused on perilipin 5 (PLIN5), an LD protein that tethers mitochondria, to probe the function and regulation of LD-mitochondria contacts. We demonstrate that efficient LD-to-mitochondria fatty acid (FA) trafficking and ß-oxidation during starvation of myoblasts are promoted by phosphorylation of PLIN5 and require an intact PLIN5 mitochondrial tethering domain. Using human and murine cells, we further identified the acyl-CoA synthetase, FATP4 (ACSVL4), as a mitochondrial interactor of PLIN5. The C-terminal domains of PLIN5 and FATP4 constitute a minimal protein interaction capable of inducing organelle contacts. Our work suggests that starvation leads to phosphorylation of PLIN5, lipolysis, and subsequent channeling of FAs from LDs to FATP4 on mitochondria for conversion to fatty-acyl-CoAs and subsequent oxidation.
    Keywords:  FATP4; PLIN5; acyl-CoA; fatty acids; lipid droplets; membrane contact sites; metabolism; mitochondria; organelles
  20. Biochim Biophys Acta Biomembr. 2023 Jun 05. pii: S0005-2736(23)00065-2. [Epub ahead of print] 184183
      In order to determine the share of protonophoric activity in the uncoupling action of lipophilic cations a number of analogues of butyltriphenylphosphonium with substitutions in phenyl rings (C4TPP-X) were studied on isolated rat liver mitochondria and model lipid membranes. An increase in the rate of respiration and a decrease in the membrane potential of isolated mitochondria were observed for all the studied cations, the efficiency of these processes was significantly enhanced in the presence of fatty acids and correlated with the octanol-water partition coefficient of the cations. The ability of C4TPP-X cations to induce proton transport across the lipid membrane of liposomes loaded with a pH-sensitive fluorescent dye increased also with their lipophilicity and depended on the presence of palmitic acid in the liposome membrane. Of all the cations, only butyl[tri(3,5-dimethylphenyl)]phosphonium (C4TPP-diMe) was able to induce proton transport by the mechanism of formation of a cation-fatty acid ion pair on planar bilayer lipid membranes and liposomes. The rate of oxygen consumption by mitochondria in the presence of C4TPP-diMe increased to the maximum values corresponding to conventional uncouplers; for all other cations the maximum uncoupling rates were significantly lower. We assume that the studied cations of the C4TPP-X series, with the exception of C4TPP-diMe at low concentrations, cause nonspecific leak of ions through lipid model and biological membranes which is significantly enhanced in the presence of fatty acids.
    Keywords:  Butyltriphenylphosphonium; Fatty acids; Mitochondria uncoupling; Nonspecific leakage; Proton transport; Pyranine-loaded liposomes
  21. bioRxiv. 2023 May 24. pii: 2023.05.24.541452. [Epub ahead of print]
      The loss of E-cadherin (E-cad), an epithelial cell adhesion molecule, has been implicated in the epithelial-mesenchymal transition (EMT), promoting invasion and migration of cancer cells and, consequently, metastasis. However, recent studies have demonstrated that E-cad supports the survival and proliferation of metastatic cancer cells, suggesting that our understanding of E-cad in metastasis is far from comprehensive. Here, we report that E-cad upregulates the de novo serine synthesis pathway (SSP) in breast cancer cells. The SSP provides metabolic precursors for biosynthesis and resistance to oxidative stress, critically beneficial for E-cad-positive breast cancer cells to achieve faster tumor growth and more metastases. Inhibition of PHGDH, a rate- limiting enzyme in the SSP, significantly and specifically hampered the proliferation of E-cad- positive breast cancer cells and rendered them vulnerable to oxidative stress, inhibiting their metastatic potential. Our findings reveal that E-cad adhesion molecule significantly reprograms cellular metabolism, promoting tumor growth and metastasis of breast cancers.
  22. EMBO Rep. 2023 Jun 05. e56430
      Human Tim8a and Tim8b are paralogous intermembrane space proteins of the small TIM chaperone family. Yeast small TIMs function in the trafficking of proteins to the outer and inner mitochondrial membranes. This putative import function for hTim8a and hTim8b has been challenged in human models, but their precise molecular function(s) remains undefined. Likewise, the necessity for human cells to encode two Tim8 proteins and whether any potential redundancy exists is unclear. We demonstrate that hTim8a and hTim8b function in the assembly of cytochrome c oxidase (Complex IV). Using affinity enrichment mass spectrometry, we define the interaction network of hTim8a, hTim8b and hTim13, identifying subunits and assembly factors of the Complex IV COX2 module. hTim8-deficient cells have a COX2 and COX3 module defect and exhibit an accumulation of the Complex IV S2 subcomplex. These data suggest that hTim8a and hTim8b function in assembly of Complex IV via interactions with intermediate-assembly subcomplexes. We propose that hTim8-hTim13 complexes are auxiliary assembly factors involved in the formation of the Complex IV S3 subcomplex during assembly of mature Complex IV.
    Keywords:  Complex IV; mitochondria; protein assembly; protein trafficking; small TIMs
  23. Exp Cell Res. 2023 Jun 03. pii: S0014-4827(23)00218-5. [Epub ahead of print] 113671
      Primary cilia (PCs) that are present in most human cells and perform sensory function or signal transduction are lost in many solid tumors. Previously, we identified VDAC1, best known to regulate mitochondrial bioenergetics, to negatively regulate ciliogenesis. Here, we show that downregulation of VDAC1 in pancreatic cancer-derived Panc1 and glioblastoma-derived U-87MG cells significantly increased ciliation. Those PCs were significantly longer than the control cells. Such increased ciliation possibly inhibited cell cycle, which contributed to reduced proliferation of these cells. VDAC1-depletion also led to longer PCs in quiescent RPE1 cells. Therefore, serum-induced PC disassembly was slower in VDAC1-depleted RPE1 cells. Overall, this study reiterates the importance of VDAC1 in modulating tumorigenesis, due to its novel role in regulating PC disassembly and cilia length.
    Keywords:  Cell cycle; Cilia disassembly; Mitochondria; Primary cilia; Proliferation; Tumorigenesis; VDAC1
  24. Cell Calcium. 2023 Jun 02. pii: S0143-4160(23)00077-5. [Epub ahead of print]113 102765
      The mitochondrial inner boundary membrane harbors a protein called MICU1, which is sensitive to Ca2+ and binds to the MICOS components Mic60 and CHCHD2. Changes in the mitochondrial cristae junction structure and organization in MICU1-/- cells lead to increased cytochrome c release, membrane potential rearrangement, and changes in mitochondrial Ca2+ uptake dynamics. These findings shed new light on the multifaceted role of MICU1, highlighting its involvement not only as an interaction partner and regulator of the MCU complex but also as a crucial determinant of mitochondrial ultrastructure and, thus, an essential player in processes initiating apoptosis.
    Keywords:  Apoptosis; Ca(2+) signaling; Cristae junction; MICOS-complex; MICU1; Mitochondria
  25. ACS Chem Biol. 2023 Jun 08.
      The crosstalk between mitochondria and the nucleus regulates cell plasticity and innate immune response. A new study shows that copper(II) accumulates in mitochondria of activated macrophages in response to pathogen infection and induces metabolic and epigenetic reprogramming that promotes inflammation. Pharmacologic targeting of mitochondrial copper(II) uncovers a new therapeutic strategy to combat aberrant inflammation and regulate cell plasticity.
  26. Blood Adv. 2023 Jun 05. pii: bloodadvances.2023009890. [Epub ahead of print]
      T cells demonstrate impaired function in Multiple Myeloma (MM), but suppressive mechanisms in the bone marrow microenvironment remain poorly defined. We observe that bone marrow CD8+ T-cell function is decreased in MM patients compared to controls, and also is consistently lower within bone marrow samples than matched peripheral blood. These changes are accompanied by decreased mitochondrial mass and markedly elevated long-chain fatty acid uptake. In vitro modelling confirmed that uptake of bone marrow lipids suppresses CD8+ T function, which is impaired in autologous bone marrow plasma, but rescued by lipid removal. Analysis of single-cell RNA-sequencing data identified expression of fatty acid transport protein 1 (FATP1) in bone marrow CD8+ T cells in MM, and FATP1 blockade also rescued CD8+ T-cell function, thereby identifying this as a novel target to augment T cell activity in MM. Finally, analysis of samples from treated patient cohorts identified that CD8+ T cell metabolic dysfunction resolves in treatment-responsive but not relapsed MM patients and is associated with substantial T cell functional restoration.
  27. J Biol Chem. 2023 Jun 01. pii: S0021-9258(23)01905-1. [Epub ahead of print] 104877
      Abcb10 is a mitochondrial membrane protein involved in hemoglobinization of red cells. Abcb10 topology and ATPase domain localization suggest it exports a substrate, likely biliverdin, out of mitochondria that is necessary for hemoglobinization. In this study we generated Abcb10 deletion cell lines in both mouse murine erythroleukemia (MEL) and human erythroid precursor human myelogenous leukemia (K562) cells to better understand the consequences of Abcb10 loss. Loss of Abcb10 resulted in an inability to hemoglobinize upon differentiation in both K562 and MEL cells with reduced heme and intermediate porphyrins and decreased levels of aminolevulinic acid synthase 2 activity. Metabolomic and transcriptional analyses revealed that Abcb10 loss gave rise to decreased cellular arginine levels, increased transcripts for cationic and neutral amino acid transporters with reduced levels of the citrulline to arginine converting enzymes argininosuccinate synthetase and argininosuccinate lyase. The reduced arginine levels in Abcb10 null cells gave rise to decreased proliferative capacity. Arginine supplementation improved both Abcb10 null proliferation and hemoglobinization upon differentiation. Abcb10 null cells showed increased phosphorylation of Eukaryotic Translation Initiation Factor 2 Subunit Alpha (eIF2A), increased expression of nutrient sensing transcription factor ATF4 and downstream targets DNA damage inducible transcript 3 (Chop), ChaC glutathione specific gamma-glutamylcyclotransferase 1 (Chac1) and arginyl-tRNA synthetase 1 (Rars). These results suggest that when the Abcb10 substrate is trapped in the mitochondria, the nutrient sensing machinery is turned on remodeling transcription to block protein synthesis necessary for proliferation and hemoglobin biosynthesis in erythroid models.
    Keywords:  Arginine; differentiation; erythroid; metabolism; nutrient; transporter
  28. Cell Rep. 2023 Jun 03. pii: S2211-1247(23)00612-5. [Epub ahead of print]42(6): 112601
      Acidic environments reduce the intracellular pH (pHi) of most cells to levels that are sub-optimal for growth and cellular functions. Yet, cancers maintain an alkaline cytoplasm despite low extracellular pH (pHe). Raised pHi is thought to be beneficial for tumor progression and invasiveness. However, the transport mechanisms underpinning this adaptation have not been studied systematically. Here, we characterize the pHe-pHi relationship in 66 colorectal cancer cell lines and identify the acid-loading anion exchanger 2 (AE2, SLC4A2) as a regulator of resting pHi. Cells adapt to chronic extracellular acidosis by degrading AE2 protein, which raises pHi and reduces acid sensitivity of growth. Acidity inhibits mTOR signaling, which stimulates lysosomal function and AE2 degradation, a process reversed by bafilomycin A1. We identify AE2 degradation as a mechanism for maintaining a conducive pHi in tumors. As an adaptive mechanism, inhibiting lysosomal degradation of AE2 is a potential therapeutic target.
    Keywords:  CP: Cancer; CP: Metabolism; acid adaptation; acid-base; acidosis; chloride/bicarbonate exchanger; colorectal cancer; intracellular pH; lysosomes; tumor acidity; tumor microenvironment
  29. J Control Release. 2023 Jun 06. pii: S0168-3659(23)00374-7. [Epub ahead of print]
      Mitochondrion is an ideal target for amplifying ROS attack in antitumor treatment. Benefiting from distinctive properties of mitochondria, the precise delivery of ROS generator to mitochondria could maximumly utilize ROS for oxidation therapy. Herein, we prepared an innovative ROS-activatable nanoprodrug (HTCF) which dually targets tumor cells and mitochondria for antitumor therapy. Cinnamaldehyde (CA) was conjugated to ferrocene (Fc) and triphenylphosphine by thioacetal linker, to synthesize mitochondria-targeting ROS-activated prodrug (TPP-CA-Fc), which subsequently self-assembled into nanoprodrug via host-guest interactions between TPP-CA-Fc and cyclodextrin-decorated hyaluronic acid conjugate. Under mitochondrial high ROS condition, especially in tumor cells, HTCF selectively initiate in-situ Fenton reaction to catalyze H2O2 into highly cytotoxic •OH, ensuring maximum generation and utilization of •OH for precision CDT. Meanwhile, the mitochondrial high ROS trigger thioacetal bond cleavage and CA release. The released CA stimulate mitochondrial oxidative stress aggravation and H2O2 regeneration, which in turn react with Fc for more •OH generation, forming self-amplifying positive feedback cycle of CA release and ROS burst. With self-augmented Fenton reaction and mitochondria-specific destruction, HTCF ultimately induce intracellular ROS burst and severe mitochondrial dysfunction for amplified ROS-mediated antitumor therapy. Such an ingenious organelles-specialized nanomedicine exhibited prominent antitumor effect both in vitro and in vivo, revealing underlying perspectives to amplify tumor-specific oxidation therapy.
    Keywords:  Mitochondria targeting; Oxidation therapy; ROS-activated; Reactive oxygen species; Self-augmented
  30. Front Pharmacol. 2023 ;14 1196158
      Background: Colorectal cancer (CRC) is one of the most common malignancies causing the third highest mortality rate in the world. It is particularly urgent to explore effective therapeutic strategies to overcome this disease. We identified a novel benzothiazole derivative (BTD) that may serve as a potentially effective agent against CRC. Method: MTT assays, cell colony formation assays, EdU staining assays, flow cytometry, RNA-seq, Western blotting, and migration and invasion assays were used to examine the effects of BTD on cell proliferation, apoptosis, metastasis, and the cell cycle. The antitumor activity of BTD in vivo was investigated in a CT26 tumor-bearing mouse model. Immunohistochemistry (IHC) was performed to examine the protein expression in mouse tumors. Hematology, biochemical analysis, and H&E staining were used to assess the biosafety of BTD. Results: We observed that BTD suppressed cell proliferation and metastasis and promoted the apoptosis of tumor cells in vitro. Treatment with BTD at a tolerable dose significantly reduced tumor growth in CT26-tumor-bearing mice and appeared to be safe. Treatment of BTD induced apoptosis by increasing the generation of reactive oxygen species (ROS) and evoking the loss of mitochondrial transmembrane potential. Overall, BTD suppressed cell proliferation and metastasis, and induced apoptosis of colorectal tumor cells through the ROS-mitochondria-mediated apoptotic pathway. The preliminary proof of the antitumor activity and relative safety of BTD were validated in a mouse model. Conclusion: Our findings suggest that BTD could serve as a potentially safe and effective candidate for CRC treatment.
    Keywords:  apoptosis; benzothiazole derivative; colorectal cancer; mitochondrial intrinsic pathway; small molecular drug
  31. J Ovarian Res. 2023 Jun 05. 16(1): 108
      The mortality rate of epithelial ovarian cancer (EOC) remains the first in malignant tumors of the female reproductive system. The characteristics of rapid proliferation, extensive implanted metastasis, and treatment resistance of cancer cells require an extensive metabolism rewiring during the progression of cancer development. EOC cells satisfy their rapid proliferation through the rewiring of perception, uptake, utilization, and regulation of glucose, lipids, and amino acids. Further, complete implanted metastasis by acquiring a superior advantage in microenvironment nutrients competing. Lastly, success evolves under the treatment stress of chemotherapy and targets therapy. Understanding the above metabolic characteristics of EOCs helps to find new methods of its treatment.
  32. Science. 2023 Jun 09. 380(6649): eabn9257
      Aging is associated with changes in circulating levels of various molecules, some of which remain undefined. We find that concentrations of circulating taurine decline with aging in mice, monkeys, and humans. A reversal of this decline through taurine supplementation increased the health span (the period of healthy living) and life span in mice and health span in monkeys. Mechanistically, taurine reduced cellular senescence, protected against telomerase deficiency, suppressed mitochondrial dysfunction, decreased DNA damage, and attenuated inflammaging. In humans, lower taurine concentrations correlated with several age-related diseases and taurine concentrations increased after acute endurance exercise. Thus, taurine deficiency may be a driver of aging because its reversal increases health span in worms, rodents, and primates and life span in worms and rodents. Clinical trials in humans seem warranted to test whether taurine deficiency might drive aging in humans.
  33. Free Radic Biol Med. 2023 Jun 02. pii: S0891-5849(23)00473-2. [Epub ahead of print]205 77-89
      NAD+ and glutathione precursors are currently used as metabolic modulators for improving the metabolic conditions associated with various human diseases, including non-alcoholic fatty liver disease, neurodegenerative diseases, mitochondrial myopathy, and age-induced diabetes. Here, we performed a one-day double blinded, placebo-controlled human clinical study to assess the safety and acute effects of six different Combined Metabolic Activators (CMAs) with 1 g of different NAD+ precursors based on global metabolomics analysis. Our integrative analysis showed that the NAD+ salvage pathway is the main source for boosting the NAD+ levels with the administration of CMAs without NAD+ precursors. We observed that incorporation of nicotinamide (Nam) in the CMAs can boost the NAD+ products, followed by niacin (NA), nicotinamide riboside (NR) and nicotinamide mononucleotide (NMN), but not flush free niacin (FFN). In addition, the NA administration led to a flushing reaction, accompanied by decreased phospholipids and increased bilirubin and bilirubin derivatives, which could be potentially risky. In conclusion, this study provided a plasma metabolomic landscape of different CMA formulations, and proposed that CMAs with Nam, NMN as well as NR can be administered for boosting NAD+ levels to improve altered metabolic conditions.
    Keywords:  Carnitine; Cysteine; Metabolomics; NAD(+) precursors; Serine; Systems medicine
  34. Clin Cancer Res. 2023 Jun 08. pii: CCR-23-0375. [Epub ahead of print]
      PURPOSE: Therapeutic resistance to frontline therapy develops rapidly in small cell lung cancer (SCLC). Treatment options are also limited by the lack of targetable driver mutations. Therefore, there is an unmet need for developing better therapeutic strategies and biomarkers of response. AURKB inhibition exploits an inherent genomic vulnerability in SCLC and is a promising therapeutic approach. Here, we identify biomarkers of response and develop rational combinations with AURKB inhibition to improve treatment efficacy.EXPERIMENTAL DESIGN: Selective AURKB inhibitor AZD2811 was profiled in a large panel of SCLC cell lines (n=57) and patient-derived xenograft (PDX) models. Proteomic and transcriptomic profiles were analyzed to identify candidate biomarkers of response and resistance. Effects on polyploidy, DNA damage and apoptosis were measured by flow cytometry and western blotting. Rational drug combinations were validated in SCLC cell lines and PDX models.
    RESULTS: AZD2811 showed potent growth inhibitory activity in a subset of SCLC, often characterized by, but not limited to, high cMYC expression. Importantly, high BCL2 expression predicted resistance to AURKB inhibitor response in SCLC, independent of cMYC status. AZD2811-induced DNA damage and apoptosis were suppressed by high BCL2 levels, while combining AZD2811 with a BCL2 inhibitor significantly sensitized resistant models. In vivo, sustained tumor growth reduction and regression was achieved even with intermittent dosing of AZD2811 and venetoclax, an FDA approved BCL2 inhibitor.
    CONCLUSIONS: BCL2 inhibition overcomes intrinsic resistance and enhances sensitivity to AURKB inhibition in SCLC preclinical models.
  35. Life Sci. 2023 Jun 03. pii: S0024-3205(23)00461-7. [Epub ahead of print] 121827
      AIMS: In this study, we aimed to investigate previously unrecognized lipid metabolic perturbations in tamoxifen-resistant breast cancer (BC) by conducting comprehensive metabolomics and transcriptomics analysis. We identified the role of 3-hydroxy-3-methylglutary-coenzyme-A-synthase 2 (HMGCS2), a key enzyme responsible for ketogenesis, in tamoxifen-resistant BC growth.MAIN METHODS: Comprehensive metabolomics (CE-TOFMS, LC-TOFMS) and transcriptiomics analysis were performed to characterize metabolic pathways in tamoxifen-resistant BC cells. The upregulation of HMGCS2 were verified thorugh immunohistochemistry (IHC) in clinical samples obtained from patients with recurrent BC. HMGCS2 inhibitor was discovered through surface plasmon resonance analysis, enzyme assay, and additional molecular docking studies. The effect of HMGCS2 suppression on tumor growth was studied thorugh BC xenograft model, and intratumoral lipid metabolites were analyzed via MALDI-TOFMS imaging.
    KEY FINDINGS: We revealed that the level of HMGCS2 was highly elevated in both tamoxifen-resistant T47D sublines (T47D/TR) and clinical refractory tumor specimens from patients with ER+ breast cancer, who had been treated with adjuvant tamoxifen. Suppression of HMGCS2 in T47D/TR resulted in the accumulation of mitochondrial reactive oxygen species (mtROS) and apoptotic cell death. Further, we identified alphitolic acid, a triterpenoid natural product, as a novel HMGCS2-specific inhibitor that elevated mtROS levels and drastically retarded the growth of T47D/TR in in vitro and in vivo experiments.
    SIGNIFICANCE: Enhanced ketogenesis with upregulation of HMGCS2 is a potential metabolic vulnerability of tamoxifen-resistant BC that offers a new therapeutic opportunity for treating patients with ER+ BC that are refractory to tamoxifen treatment.
    Keywords:  Alphitolic acid; Breast cancer; HMGCS2; Tamoxifen; Tamoxifen resistance